Reactive Oxygen Metabolite Production is Inhibited by Histamine and H 1 -antagonist Dithiaden in Human PMN Leukocytes

The study evaluated the distinction between extracellular and intracellular production of reactive oxygen metabolites (ROM) in isolated polymorphonuclear leukocytes (PMNL) stimulated with opsonised zymosan (OZ) and investigated its modulation by the endogenous mediator histamine (0.1-100 &#119 mol/l) and by the H 1 -antagonist dithiaden (1-100 &#119 mol/l). For this observation, a modified luminol and an isoluminol amplified chemiluminescence (CL) technique were used. Our results showed that PMNL activated with OZ responded with a respiratory burst accompanied by both extra- and intracellular generation of ROM. Histamine and dithiaden significantly decreased both the extra- and intracellular component of chemilumiescence stimulated with OZ. While dithiaden decreased both the extra- and intracellular part of CL with the same potency, histamine decreased preferentially the extracellular part of CL. The fact that histamine as well as the H 1 -antagonist dithiaden decreased the respiratory burst indicates that not only histamine receptors but also non-receptor mechanisms could be involved in the reduction of CL. Interaction with enzymes (NADPH-oxidase, myeloperoxidase, phospholipase A 2 ) or interference with PMNL membrane structure may well result in reduction of the chemiluminescence signal.     

Luminol-dependent chemiluminescence in bovine eosinophils and neutrophils: Differential increase of intracellular and extracellular chemiluminescence induced by soluble stimulants

Luminol chemiluminescence was used to detect activation of the respiratory burst oxidase in bovine eosinophils and neutrophils. Extracellular and intracellular chemiluminescence were measured by supplementing the medium with horseradish peroxidase and catalase, respectively. Pure bovine eosinophils (> 90%), maximally stimulated with 1 nmol/l phorbol 12-myristate-13-acetate (PMA) showed ten times more extracellular luminol-dependent chemiluminescence (CL) than maximally stimulated pure bovine neutrophils (> 96%). Extracellular CL from eosinophils was preferably induced over intracellular CL by both PMA (27-fold difference) and platelet-activating factor (PAF) at 2 μmol/l (9-fold difference), but not by calcium ionophore A23187 (15 μmol/l). Time course information was used in the following experiments to distinguish between the mode of action of various stimulants. A progressively longer lag period was observed in eosinophil suspensions treated with decreasing doses of PMA, whereas platelet-activating factor induced a dose-dependent increase in the maximum response with no change in time to peak CL. The time course of extracellular CL was almost identical to intracellular CL for all stimulants tested, providing no evidence to suggest that extracellular CL stems from a different enzyme system than intracellular CL. Eosinophils generated most extracellular CL when stimulated with PMA, whereas neutrophils were most efficiently stimulated with A23187, which induced intracellular CL in eosinophils as well as in neutrophils. This accords with the greater tendency of neutrophils to ingest and kill microorganisms, whereas eosinophils are armed to destroy large extracellular targets.     

Inhibitory effect of stobadine on FMLP-induced chemiluminescence in human whole blood and isolated polymorphonuclear leukocytes.

The chemiluminescence (CL) technique with luminol and isoluminol was used to characterize the effect of stobadine on reactive oxygen metabolites (ROM) generation in human whole blood and in isolated polymorphonuclear leukocytes (PMNL) stimulated with N-formyl-methionyl-leucyl-phenyl-alanine (FMLP). In whole blood and in isolated PMNL, stobadine in the concentrations of 1, 10 and 100 micromol/L significantly inhibited the CL signal after FMLP, which activated predominantly extracellular generation of ROM. The same concentrations of stobadine were effective on CL in a cell-free system. On the other hand, myeloperoxidase (MPO) liberation was decreased by stobadine only in the concentration of 100 micromol/L. The results showed stobadine to act as a potent inhibitor/scavenger of extracellularly produced ROM in human PMNL and indicated interference of stobadine with ROM as well as with signalling events resulting in NADPH-oxidase activation and MPO liberation.     

Contrasting effects of Mycoplasma fermentans and M. felis on the viability and chemiluminescence response of human polymorphonuclear leukocytes

Abstract Trypan blue exclusion was used to estimate the viability of human polymorphonuclear leukocytes (PMNL) in the presence of Mycoplasma felis and two strains of M. fermentans (PG18 and incognitus). The competence of PMNL to mount a respiratory burst when challenged with the mycoplasmas was also monitored by luminol-dependent chemiluminescence (CL). Both un-opsonised and non-immune human serum opsonised M. felis cells had little effect on PMNL viability. In contrast, PMNL viability was reduced markedly by un-opsonised cells of M. fermentans strain incognitus and, to a lesser extent, strain PG18, and opsonisation of these mycoplasmas further enhanced killing. Death of PMNL in the presence of M. fermentans was not associated with the autonomous production of active oxygen species during the respiratory burst as M. felis induced a high CL response from PMNL, whereas that induced by M. fermentans strain incognitus was significantly lower. M. fermentans may invade mammalian cells and it is suggested that the mechanism of PMNL death could be related to the ability of M. fermentans to penetrate host cell membranes.     

Effect of different inhibitors on the intracellularly and extracellularly generated chemiluminescence induced by formylmethionyl-leucyl-phenylalanine in polymorphonuclear leukocytes. Cellular response in the presence of mannitol,benzoate, taurine,indomethacin and NDGA

When polymorphonuclear leukocytes (PMNL) interact with the soluble stimulus formylmethionyl-leucyl-phenylalanine (FMLP), the cells increase their production of oxidative metabolites. This increased production can be measured as lumino-amplified light emission or chemiluminescence (CL). In the present report, experimental systems which allow a quantitation of extracellularly and intracellularly generated metabolites have been used, and the effect of mannitol, benzoate, taurine, indomethacin and nordihydroguaiaretic acid has been investigated. The presence of the hypochlorous acid scavenger taurine had no effect on the intracellular response, whereas the extracellular response was reduced with around 50%. The hydroxyl radical scavenger mannitol had only minor effects on the response, whereas benzoate, another hydroxyl radical scavenger, reduced the extracellular response with around 50% and the intracellular response with more than 90%. Indomethacin, an inhibitor of arachidonic acid metabolism, did not influence the response, whereas NDGA, also an inhibitor of the arachidonic acid metabolism, totally abolished both the extracellular and the intracellular response. The use of scavengers/inhibitors as a means of determining the mechanisms of light emission, and the origin of chemiluminescence produced by neutrophils stimulated by FMLP is discussed.     

α-tocopherol modulates tyrosine phosphorylation in human neutrophils by inhibition of protein kinase C activity and activation of tyrosine phosphatases

-Tocopherol augmentation in human neutrophils was investigated for effects on neutrophil activation and tyrosine phosphorylation of proteins, through its modulation of protein kinase C (PKC) and tyrosine phosphatase activities. Incubation of neutrophils with -tocopherol succinate (TS) resulted in a dose-dependent incorporation into cell membranes, up to 2.5 nmol/2 × 10⁶ cells. A saturating dose of TS (40 μmol/l) inhibited oxidant production by neutrophils stimulated with phorbol myristate acetate (PMA) or opsonized zymosan (OZ) by 86 and 57%, as measured by luminol-amplified chemiluminescence (CL). With PMA, TS inhibited CL generation to a similar extent to staurosporine (10 nmol/l) or genistein (100 μmol/l), and much more than Trolox (40 μmol/l). With OZ, TS inhibited CL to a similar extent to Trolox. Neutrophil PKC activity was inhibited 50% or more by TS or staurosporine. The enzyme activity was unaffected by genistein or Trolox, indicating a specific interaction of -tocopherol. TS or Trolox increased protein tyrosine phosphorylation in resting neutrophils, and as with staurosporine further increased tyrosine phosphorylation in PMA-stimulated neutrophils, while the tyrosine kinase (TK) inhibitor genistein diminished phosphorylation. These effects in resting or PMA-stimulated neutrophils were unrelated to protein tyrosine phosphatase (PTP) activities, which were maintained or increased by TS or Trolox. In OZ-stimulated neutrophils, on the other hand, all four compounds inhibited the increase in tyrosine-phosphorylated proteins. In this case, the effects of pre-incubation with TS or Trolox corresponded with partial inhibition of the marked (85%) decrease in PTP activity induced by OZ. These results indicate that -tocopherol inhibits PMA-activation of human neutrophils by inhibition of PKC activity, and inhibits tyrosine phosphorylation and activation of OZ-stimulated neutrophils also through inhibition of phosphatase inactivation.     

The influence of age and sex on phagocyte chemiluminescence

The process of ageing is associated with increased susceptibility to infection. Phagocytes form the primary defence mechanism against infecting microorganisms, but the influence of ageing on phagocyte function remains controversial. In this study we have applied a microtitre plate phagocyte chemiluminescence (CL) assay suitable for clinical use to compare phagocyte oxidative metabolism in younger healthy subjects (age 20–60 years) and healthy older (60–70 years) subjects. Polymorphonuclear leukocytes (PMNL) and monocytes were stimulated using phorbol myristate acetate (PMA), serum opsonized zymosan (SOZ), and non-opsonized zymosan (ZYM) in the presence of both lucigenin and luminol. Monocytes showed a higher luminolenhanced CL response to PMA in males compared with females in the younger age group. No PMNL differences were observed between the sexes. Although no difference were found in relation to age when cells were stimulated with PMA and SOZ, significantly lower background (unstimulated) CL was obtained from PMNL with luminol. PMNL luminol-enhanced CL responses were also lower in response to ZYM. The findings suggest a reduced response of PMNL from older subjects to minimal stimulation. This could be related to abnormalities in the triggering of the respiratory burst or myeloperoxidase release due to ageing. The influence of age and sex should be taken into account in clinical studies of phagocyte CL.     

Luminol‐, isoluminol‐ and lucigenin‐enhanced chemiluminescence of rat blood phagocytes stimulated with different activators

Luminol‐, isoluminol‐ or lucigenin‐enhanced chemiluminescence (CL) was used to measure the production of reactive oxygen species by rat blood leukocytes. Opsonized zymosan (OZ), phorbol‐12‐myristate‐13‐acetate (PMA), calcium ionophore A23187 (Ca‐I) or N‐formyl‐Met‐Leu‐Phe (fMLP) were used as activators. The CL signal of isolated blood leukocytes decreased in rank order of luminol > isoluminol > lucigenin. The kinetic pro?les of luminol‐ and isoluminol‐enhanced CL were similar upon stimulation by each activator tested. The remarkably higher luminol and isoluminol CL responses were obtained after OZ stimulation when compared with other activators. However, when lucigenin was used, the PMA‐ and OZ‐stimulated CL were comparable. The presence of plasma increased OZ‐activated CL because of the enhanced phagocytosis of OZ. This was demonstrated by determining the phagocytosis of the ?uorescent OZ using a ?ow cytometer. In contrast, the presence of plasma decreased PMA‐activated CL, due to the antioxidant properties of plasma as determined by the CL method. As far as whole blood is concerned, only OZ activated luminol‐enhanced CL was reliable. Blood volumes over 5 µL decreased CL activity due to the scavenging ability of erythrocytes. The results suggest that 0.5 µL whole blood is suf?cient for routine luminol‐enhanced CL analysis of whole blood oxidative burst in rats. Copyright © 2004 John Wiley & Sons, Ltd.     

Automation and computerization of chemiluminescence: A new methodological approach in the study of human phagocytes

Peripheral blood phagocytic cells (PMNLs) are activated by contact with opsonized particles. Metabolic activation of PMNLs is associated with a remarkable increase in the respiratory burst and generates high energy oxygen compounds which are responsible for the bactericidal activity of PMNLs and for their ability to produce luminol-dependent chemiluminescence (CL). The CL phenomenon is measured by an automated and computerized photoluminometer (Berthold LB950) in whole blood stimulated with opsonized zymosan. This whole blood method of CL measurement has been applied to the study of the phagocytic process and to the investigation of cellular and humoral abnormalities in several pathologies, indicating this assay as a simple, rapid and reliable test.     

Localization of the luminol-dependent chemiluminescence reaction in human granulocytes

The granulocyte luminol-dependent chemiluminescence (CL) reaction is linked to the enzyme myeloperoxidase reacting with products of the respiratory burst activation. The results presented in this paper, show that the light generated in granulocytes originate both from intracellular and extracellular reactions; however, depending on the stimulus used the one or the other will dominate the activity measured. Furthermore, lysosomal fusion is proposed to be required for the intracellular CL reaction.     

Palmitic Acid Anilide-Induced Respiratory Burst In Human Polymorphonuclear Leukocytes is Inhibited by a Protein Kinase C Inhibitor, Ro 31-8220

Human polymorphonuclear leukocytes (PMNL) were exposed to palmitic acid anilide, an impurity in the case oils that caused the Spanish Toxic Oil Syndrome in 1981, and to the corresponding fatty acid, palmitic acid. The effects of these compounds were studied on the production of reactive oxygen metabolites (ROM) and changes in the levels of free intracellular calcium. Palmitic acid anilide induced the production of reactive oxygen metabolites in PMNL. Interestingly, the palmitic acid anilide-induced respiratory burst was completely blocked by a protein kinase C inhibitor, Ro 31-8220. Moreover, palmitic acid anilide additively amplified the production of ROM caused by a chemotactic peptide, formyl-Methionyl-Leucyl-Phenylalanine (FMLP). In contrast, palmitic acid anilide did not have any effect on the production of ROM induced by a tumor promoter, phorbol myristate acetate (PMA). Palmitic acid, in turn, did not markedly induce the production of ROM nor did it amplify the agonist-induced respiratory burst. Neither of the compounds, alone or in combination with FMLP, affected the levels of intracellular calcium in PMNL. These results indicate that the aniline moiety in palmitic acid modifies its effects on the activation of human PMNL, and the subsequent oxidative burst. The present results also suggest that palmitic acid anilide may activate PMNL through a protein kinase C-dependent mechanism. © 1997 Elsevier Science Inc.     

Cell-Dependent Chemiluminescence. Modulation of the N-Formyl Chemotactic Peptide (Fnlpntl) Mediated Oxidative Burst in Human Polymorphonuclear Leukocytes (Pmnl) By Murine Monoclonal Antibody Nms-1

Binding of purified monoclonal antibody (moAB) IgM NMS-1 to suspended initially spherical living human PMNLs is not associated with the generation of chemiluminescence but was found to enhance the chemiluminescence response to the N-formyl chemotactic peptide FNLPNTL.

We investigated quantitatively the kinetics of oxygen metabolite generation by PMNLs stimulated with FNLPNTL ± moAB NMS-1 using luminol-dependent chemiluminescence as a very sensitive detection system. Chemiluminescence detection allowed the analysis of the time sequence of onset and development of reactive oxygen metabolites following stimulation of PMNLs by FNLPNTL in the presence of moAB NMS-1. The increase of response of PMNLs stimulated with FNLPNTL in the presence of moAB NMS-1 depended on the concentration of the antibody and the sequence of stimulus addition.

Stimulation of human PMNLs by 10nM FNLPNTL induced a rapid burst of chemiluminescence which peaked ∼5min after stimulus addition. The subsequent addition of moAB NMS-1 (≥2μg/ml DPBS(+)—0.1% HSA, 37°C) to FNLPNTL-stimulated PMNLs—after the FNLPNTL-mediated response had already decayed (16-18 min) - without delay induced a second burst of oxygen metabolite generation. The magnitude of this second peak of activation was dose-dependent.

Treatment of PMNLs with moAB NMS-1 (≥ 1μg/ml DPBS(+)—0.1% HSA, 3 min, 37°C)—prior to FNLPNTL (10nM) stimulation - increased rate and magnitude of the FNLPNTL-mediated response. This response is biphasic with the first peak at the FNLPNTL position and a second, higher peak ∼16 min after FNLPNTL addition. The magnitude of response was dose-dependent. The latency (lag time) of the respone was not changed compared to controls which received no moAB NMS-1 treatment.

The observed moAB NMS-1 dependent increase in FNLPNTL-mediated chemiluminescence is transient (5-60 min), persistent activation was not detected.     

Model components of luminol chemiluminescence generated by PMNL

The production of activated oxygen species (AOS) by neutrophils (PMNL) is thought to play a key role in the host defence against invading microorganisms. However, the oxygen metabolites are toxic not only to the invading bacteria but also to the surrounding tissue. The oxidative metabolites production can be evaluated by means of chemiluminescent methods. In this study, the possibility of a new analytical approach for quantitative assessment of chemiluminescent kinetics (AOS generation) of isolated PMNL was estimated.

Based on the assumption that the kinetics of luminol-amplified chemiluminescence (LCL) of stimulated PMNL possesses a time-probabilistic nature, this kinetics was described with three components. These components, obtained from different investigated systems, were analyzed and a conclusion was made that the first and the second component represent the processes resulting in extra-and intracellular myeloperoxidase (MPO)-dependent light emission (AOS generation), respectively. The second component was found to be completely dependent on the stimulus ingestion. The third component was not completely MPO-dependent and complicated for interpretation. This component was weakly dependent on the stimulus ingestion, and presents at least some intracellular processes different from those presented by the second component.

A conclusion is made that the examined approach for analysis of LCL kinetics allows an assessment of extra-and intracellularly generated quantities of AOS by stimulated PMNL. The assessment could be done for emitting systems in which no additional modificators are used.     

Inhibitory and enhancing effects of piroxicam on whole blood chemiluminescence.

The effects of piroxicam on the production of reactive oxygen species by stimulated phagocytes was studied in whole blood by a chemiluminescence (CL) technique in relation to maximum activity, localization and kinetics of radical generation. We found that piroxicam dose-dependently inhibited total (intra- and extracellular) zymosan-stimulated luminol CL (LCL) at a high stimulant concentration (p = 0.0001). Piroxicam additionally decreased cytochalasin B-reduced LCL, which shows that the effect of the drug should be sought in the extracellular component of the response. Piroxicam inhibited the first phase of extracellular LCL in a dose-dependent manner (p = 0.0001) and revealed itself as an enhancing agent of CL in later time intervals after the start of respiratory burst, in a model system containing horseradish peroxidase (HRP) and sodium azide. It enhanced LCL of a cell-free system, i.e. influenced the CL due to HRP-catalysed decomposition of hydrogen peroxide. It also dose-dependently inhibited the early extracellular superoxide production, evaluated by lucigenin CL (p = 0.022). Piroxicam inhibited the total fMLP-stimulated LCL by 70% approximately and, only by about 30%, the first phase of fMLP-stimulated extracellular LCL, which presupposes an effect on myeloperoxidase-catalysed formation of hypochloric acid. Piroxicam slightly increased the intracellular LCL by phagocytes (p = 0.02), an effect that is probably connected with its ability to induce the release of secondary messengers in signal transduction. In conclusion, the anti-inflammatory effect of piroxicam is probably related to the inhibition of the extracellular generation of superoxide and hypochloric acid in the early stages of phagocyte activation.     

Temperature influence on stimulated PMN respiratory burst.

Body temperature can modulate the pathogenesis of infectious, metabolic and autoimmune diseases. This effect has been attributed to several hypothesized mechanisms. Body temperature could play an important role in influencing some cellular functions of human white blood cells. In this work we examined the temperature effect on the respiratory burst in human neutrophils. Human polymorphonuclear leucocytes (PMN) were obtained from heparinized venous blood by dextran sedimentation and erythrocyte lysis with NH4Cl (0.87%). Granulocytes were stimulated with opsonized zymosan (OZ), formyl-methionyl-leucyl-phenylalanine (FMLP), phorbol myristate acetate (PMA), and monosodium urate (MSU) crystals at different temperatures (26, 37, 39, 40, 42 degrees C). The technique of luminol dependent chemiluminescence (CL) was used as indicator of oxygen free radicals (OFR) release by stimulated cells. OFR production from PMN stimulated with OZ, PMA, FMLP was higher at 37 degrees C than at 26, 39, 40, 42 degrees C (p < 0.001 OZ stimulated PMN at 40-42 degrees C; p < 0.05 PMA stimulated PMN at 42 degrees C. Significantly different from 37 degrees C value). OFR release from PMN stimulated with MSU crystals was significantly increased at 39 degrees C compared to 37 degrees C value (p < 0.001). This effect could not only be attributed to temperature influence on neutrophil activity. The specific polymorphonuclear leukocyte response to the microcrystals and the temperature influence on chemical and physical characteristics of the crystals may play an important role. We are now studying the temperature effect on activity of PMN exposed to others crystals.     

Factors affecting the measurement of chemiluminescence in stimulated human polymorphonuclear leucocytes

Optimum conditions were established for the generation and measurement of luminoldependent chemiluminescence (CL) in human polymorphonuclear leucocytes (PMNL) stimulated with a variety of particulate and soluble agents. Several factors had a particular influence on the kinetics of CL stimulated by the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP). Two peaks, both azide-sensitive, were observed at 21°C and 25°C. but these increased in magnitude and merged t o give a single, early peak when the temperature was increased t o 37°C. Pre-exposure of PMNL to a buffer containing calcium was essential for the expression of both phases of fMLP-stimulated CL, while the second peak decreased dramatically if the cells were stored at 4°C for 4 hours before assay. In contrast, storage of PMNL at 4°C for up t o 8 hours in a buffer without divalent cations did not alter the kinetics or magnitude of CL induced by other stimuli, and had the benefit of minimizing the rate of cell aggregation. This study confirms that measurement of luminol-dependent CL in stimulated PMNL is a useful analytical tool, but shows that careful attention t o experimental design is required t o ensure that the observed CL provides a true measure of the parameter under investigation.     

Upregulation of Respiratory Burst of Polymorphonuclear Leukocytes By A Bleomycin Derivative, Peplomycin

The influence of peplomycin (PLM) on the respiratory burst of peripheral blood polymorphonuclear leukocytes (PMN) was investigated. Short-term (5 min) treatment of human PMN with 0.1μg/ml to 100μg/ml of PLM increased phorbol myristate acetate (PMA)-and formyl-methionyl-leucyl-phenylalanine (FMLP)-induced luminol-dependent chemiluminescence. PMN, as well as alveolar macrophages from rabbits treated with 0.5 to 1.0 mg/kg of peplomycin per day for 5 days, generated more superoxide (O₂^-) than the cells from untreated rabbits. In both PLM-treated and untreated PMN, chemiluminescence induced by FMLP and PMA was decreased to less than 50% of the control by staurosporine, superoxide dismutase (SOD) and catalase. However, the peak intensity in PLM-untrcated PMN was decreased to about 30% of the control by genislein, while this agent induced a slight decrease in peak intensity in the PLM-treated PMN. Inositol triphosphate and diacyl glycerol levels were not clearly increased by PLM, but an increase of intracellular Ca and a shift of protein kinase C (PKC) to the membrane occurred in PMN within 1 min after PLM treatment. Western blotting revealed that the tyrosine phosphorylation of a 115 kDa protein was upregulated by 5 to 50μg/ml of PLM. While, PLM suppressed SOD activity in alveolar macrophages and PMN. These results seem to indicate that PLM increases the respiratory burst of PMN and macrophages both by way of direct PKC activation and by the upregulation of protein tyrosine phosphorylation. This increased reactive oxygen generation, together with the suppression of SOD activity seems to be tissue-impairing.     

Lipopolysaccharide-dependent enhancement of adherence-mediated chemiluminescence response of polymorphonuclear leukocytes

Adherence of resting polymorphonuclear leukocytes to nylon fibre increased the chemiluminescence response (CL) from 99400 to 910300 cpm/25000 PMNL. This effect could be amplified by lipopolysaccharide (LPS) priming of granulocytes in a dose-dependent fashion. The results of nylon fibre adherence experiments suggest an in vitro model that might approximate certain conditions of in vivo PMNL-endothelial adherence and respiratory burst activation, and these reactions of polymorphonuclear leukocytes may contribute to the pathomechanisms of the Adult Respiratory Distress Syndrome.     

Sesquiterpene lactone from Wunderlichia crulsiana inhibits the respiratory burst of leukocytes triggered by distinct biochemical pathways

The sesquiterpene lactone tubiferin was chemically purified from the brazilian native plant Wunderlichia crulsiana and identified by NMR and GC/MS data. Its ability to inhibit the respiratory burst of peritoneal inflammatory polymorphonuclear leukocytes (PMN) stimulated upon addition of phorbol miristate acetate (PMA), opsonized zymosan (OZ), and N-formyl-methionyl-leucyl-phenylalanine (fMLP) was evaluated. The tubiferin inhibition was more pronounced when PMN were stimulated through the protein kinase C pathway (PMA) compared to the alternative complement pathway (OZ). The inhibition when PMN were triggered by a chemoattractant stimulus (fMLP) was similar to that achieved with OZ-stimulated phagocytes. Tubiferin showed dose-dependent effects on the PMN respiratory burst triggered by the three different substances, and also decreased substantially the carrageenan-induced mice paw edema.     

The levels of neutrophils oxidative burst in rheumatic disorders

We investigated the participation of the polymorphonuclear leukocytes (PMNs) as mediators of tissue destruction in 2 variants of rheumatic disorders that affect primarily the joints: the rheumatoid arthritis (RA) and the osteoarthritis (OA). We noted significant differences in the number of PMNs present at the joint level, which was low in OA and high in RA. The unstimulated and in vitro stimulated with opsonized zymozan (OZ) or ConA PMNs from the peripheral blood of these two groups of patients released a normal level of oxygen free radicals (OFR). On the contrary, the neutrophils isolated from the synovial fluid (SF) of RA patients showed a distinct feature: the unstimulated cells released high amounts of OFR, while those in vitro stimulated with OZ or ConA presented a low level of respiratory burst. In OA both in vitro unstimulated and stimulated with OZ or ConA neutrophils isolated from the SF had diminished values of OFR released. We assume that the chemiluminescence assay could detect distinct differences of the PMNs implications in RA and OA injuries and may be a laboratory test in the differential diagnosis of these rheumatic disorders.